Human GRK5 Baculovirus-Insect cells Overexpression Lysate

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Human GRK5 Baculovirus-Insect cells Overexpression Lysate: Product Information

Product Description
This Human GRK5 overexpression lysate was created in Baculovirus-Insect cells and intented for use as a Western blot (WB) positive control. Purification of GRK5 protein (Cat: 10839-H08B) from the overexpression lysate was verified.
Expression Host
Baculovirus-Insect cells
Species
Human
Sequence Information
A DNA sequence encoding the human GRK5 (NP_005299.1) (Met 1-Ser 590) was fused with a polyhistidine tag at the C-terminus.
Molecule Mass
The recombinant human GRK5 consists of 600 amino acids and predicts a molecular mass of 69 kDa. The apparent molecular mass of rhGRK5 is approximately 58 kDa in SDS-PAGE under reducing conditions.

Human GRK5 Baculovirus-Insect cells Overexpression Lysate: Usage Guide

Preparation Method
Cell lysate was prepared by homogenization of the over-expressed cells in ice-cold modified RIPA Lysis Buffer with cocktail of protease inhibitors (Sigma). Cell debris was removed by centrifugation. Protein concentration was determined by Bradford assay (Bio-Rad protein assay, Microplate Standard assay). The cell lysate was boiled for 5 min in 1 x SDS loading buffer (50 mM Tris-HCl pH 6.8, 12.5% glycerol, 1% sodium dodecylsulfate, 0.01% bromophenol blue) containing 5% b-mercaptoethanol, and lyophilized.
Lysis Buffer
Modified RIPA Lysis Buffer: 50 mM Tris-HCl pH 7.4, 150 mM NaCl, 1mM EDTA, 1% Triton X-100, 0.1% SDS, 1% Sodium deoxycholate, 1mM PMSF.
Recommend Usage
1.  Centrifuge the tube for a few seconds and ensure the pellet at the bottom of the tube. 2.  Re-dissolve the pellet using 200μL pure water and boil for 2-5 min.
Sample Buffer
1 X Sample Buffer (1 X modified RIPA buffer+1 X SDS loading buffer).
Stability & Storage
Store at 4℃ for up to twelve months from date of receipt. After re-dissolution, aliquot and store at -80℃ for up to twelve months. Avoid repeated freeze-thaw cycles.
Application
Western Blot (WB)
Optimal dilutions/concentrations should be determined by the end user.

Human GRK5 Baculovirus-Insect cells Overexpression Lysate: Alternative Names

Human GPRK5 Overexpression Lysate

GRK5 Background Information

G protein-coupled receptor kinase 5, also known as G protein-coupled receptor kinase GRK5 and GRK5, is a member of the protein kinase superfamily, AGC Ser/Thr protein kinase family and GPRK subfamily. GRKs specifically phosphorylate agonist-occupied G protein-coupled receptors at the inner surface of the plasma membrane (PM), leading to receptor desensitization. GRKs utilize a variety of mechanisms to bind tightly, and sometimes reversibly, to cellular membranes. GRKs play an important role in mediating agonist-specific desensitization of numerous G protein-coupled receptors.
GRK5 contains oneAGC-kinase C-terminal domain, oneprotein kinase domain and oneRGS domain. GRK5 specifically phosphorylates the activated forms of G protein-coupled receptors. Phospholipid-stimulated autophosphorylation may represent a novel mechanism for membrane association and regulation of GRK5 activity. GRK5 deficiency significantly exaggerates microgliosis and astrogliosis in the presence of an inflammatory initiator, such as the excess fibrillar Abeta and the subsequent active inflammatory reactions. GRK5 deficiency has been linked to early Alzheimer's disease in humans and mouse models of the disease.
Full Name
G protein-coupled receptor kinase 5
References
  • Kunapuli,P. et al., 1994, J Biol Chem. 269 (14):10209-12.
  • Millman,E.E. et al., 2004, Br J Pharmacol  141 (2):277-84.
  • Thiyagarajan,M.M. et al., 2004, J Biol Chem  279 (17):17989-95.
  • Suo,Z. et al., 2007,Neurobiol Aging. 28 (12):1873-88.
  • Li,L. et al., 2008,J Neuroinflammation. 5 :24.
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